Cell Agglutination Research Articles

Novel Selective Galectin-3 Antagonists Are Cytotoxic to Acute Lymphoblastic Leukemia

Galectin-3 is a β-galactoside-specific, carbohydrate-recognizing protein (lectin) that is strongly implicated in cancer development, metastasis, and drug resistance. Galectin-3 promotes migration and ability to withstand drug treatment of B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cells. Due to high amino acid conservation among galectins and the shallow nature of their glycan-binding site, the design of selective potent antagonists targeting galectin-3 is challenging. Herein, we report the design and synthesis of novel taloside-based antagonists of galectin-3 with enhanced affinity and selectivity. The molecules were optimized by in silico docking, selectivity was established against four galectins, and the binding modes were confirmed by elucidation of X-ray crystal structures. Critically, the specific inhibition of galectin-3-induced BCP-ALL cell agglutination was demonstrated. The compounds decreased the viability of ALL cells even when grown in the presence of protective stromal cells. We conclude that these compounds are promising leads for therapeutics, targeting the tumor-supportive activities of galectin-3 in cancer.

Screening of mushrooms from the woodlands of Zimbabwe: Occurrence of lectins and partial purification of a mucin specific lectin from Boletus edulis.

Mushrooms are known to possess a diversity of bioactive compounds that include lectins, which are proteins or glycoproteins that bind specifically to cell surface carbohydrates, culminating in cell agglutination. The present study describes the screening of lectin activity from ten local mushrooms, namely, Amanita zambiana, Boletus edulis, Cantharellus heinemannianus, Cantharellus miomboensis, Cantharellus symoensii, Lactarius kabansus, Amanita sp., Coprinus sp., Ganoderma lucidum and Trametes strumosa. The lectin content was detected by the haemagglutination activity of mushrooms against sheep and goat erythrocytes. Among the different mushrooms screened Amanita sp., Boletus edulis and Lactarius kabansus showed high lectin activity (39, 617 and 77 HAU/mg mushroom, respectively). Boletus edulis was used for the haemagglutination inhibition assay. A total of twenty sugars and sugar derivatives, namely, α-lactose, D-glucose, D-mannose, D-raffinose, N-acetyl glucosamine, maltose, melibiose, D-ribose, porcine mucin, D-cellobiose, D-arabinose, α-methyl-D-glucoside, methyl-α-D-mannopyranoside, D-trehalose, L-arabinose, L-sorbose, L-lyxose, β-lactose, DL-xylose, and D-galactose, were used for the haemagglutination inhibition assay. Of the various carbohydrates tested, only porcine mucin was found to be the most potent inhibitor of Boletus lectin. The lectin from Boletus mushroom was partially purified using ammonium sulphate precipitation. The highest lectin activity was observed in the 30%-60% fraction. This study revealed for the first time the occurrence of lectins in the local Zimbabwean mushrooms studied as well as isolation of a novel mucin-specific lectin. The information obtained can be used for further investigation of cell surface sugars, purification and characterisation of glycoproteins and their contribution towards the medicinal properties of local mushrooms.

Increased Mean Cell Hemoglobin Concentration.

A 44-year-old man with a history of Grave disease presented with tachycardia, weight loss, and dysphagia. He was diagnosed with severe hyperthyroidism, diabetic ketoacidosis, and hyponatremia (Table 1). A complete blood count was performed and the hemoglobin, mean cell hemoglobin (MCH), and MCH concentration (MCHC) were flagged for review due to MCHC >37.5 g/dL. Smear review was normal, and the sample was prewarmed at 37°C with similar results. ... What are potential causes of an increased MCHC? Why does a markedly increased MCHC result serve as a flag for automated hematology analyzers? What is the most likely cause of the increased MCHC in this patient? The answers are below. Causes of high MCHC include red blood cell agglutination, hemolysis, lipemia, icterus, leukocytosis, spherocytosis, and electrolyte abnormalities (1). The high MCHC flag serves to detect interference in hemoglobin, hematocrit, mean cell volume, MCH, and MCHC. Hyponatremia can cause an artificially high MCHC and this was suspected in this patient. Hyponatremia can cause in vitro shrinking of the red blood cells due to osmolality differences between patient plasma and reagent diluent, leading to a falsely low hematocrit, low mean cell volume, and high MCHC (2–4).

Histological and Physiological assessment of Endothelin-1 and Cholesterol in breast cancer of women

The current study included two aspects, one of which is histological and the other physiological. It included (75) blood samples from women with breast cancer, in addition to (15) blood samples for healthy women as control, in addition to collecting tissue biopsy from the mammary glands of some women whose blood samples were drawn Including them in advance. Those diagnosed with breast cancer and ages ranged between 38-68 years, where the study was conducted from 1 May 2019 to 2 October 2019. Gland tissue and the owner of those tumours swelling of the axillary lymph nodes. Another cancer that was found was papillary carcinoma, where epithelial hyperplasia appeared in the form of large papillae that filled the cavity of the gland. There is a large number of white blood cells and severe hematoma. Another type of gland cancer discovered is cribriform carcinoma, which is a rare case. Invasive cancer is the most dangerous, as it is observed in the glandular tissue with the presence of agglutination and crowding of tumour cells and tumour cysts. The study also showed haematological changes characterized by a highly significant concentration of Endothelin-1 (P< 0.01) in the blood of women with breast cancer group 5 and group 4, compared to the control group as a usual case. At the same time, there was a significant increase (P < 0.05) in the con- centration of Endothelin-1 in infected women in groups 3, 2 and 6 compared to the control group. The study showed a very significant decrease (P < 0.01) in the cholesterol concentration of women with breast cancer of group 5, and group 6 compared to the control group, while its level decreased significantly (P ≤ 0.05) in-group 4 compared to the control group.

Glycosylation-Dependent Induction of Programmed Cell Death in Murine Adenocarcinoma Cells.

Altered surface glycosylation is a major hallmark of tumor cells associated with aggressive phenotype and poor prognosis. By recognizing specific carbohydrate motifs, lectins can be applied to distinguish tumor from healthy cells based on the expression of glycosylation-dependent markers. Through their ability to bind to specific carbohydrates, lectins induce cell agglutination and cross-link surface glycoproteins, thereby mediating mitogenic and death-inducing effects in various cell types. The carbohydrate-selective cytotoxic effect of lectins also enables their possible application in therapies targeting cancer cells. To clarify the intracellular pathways mediating cell death induced by a group of plant and fungal lectins, we investigated mouse adenocarcinoma MC-38 cells harboring inactive genes involved in apoptosis, necroptosis and pyroptosis. Treatment of MC-38 cells with wheat germ agglutinin, Maackia amurensis lectin I, and Aleuria aurantia lectin induced multiple cell death pathways through reactions that relied on the autophagy machinery without depending on caspase activation. Furthermore, inhibition of de novo protein synthesis by cycloheximide strongly decreased the cytotoxic response, indicating that the lectins investigated induced cell death via effector molecules that are not expressed under normal circumstances and supporting the non-apoptotic nature of cell death. The broad cytotoxic response to lectins can be beneficial for the development of combination therapies targeting tumor cells. Given that tumors acquire resistance to various cytotoxic treatments because of mutations in cell death pathways, compounds inducing broad cytotoxic responses, such as lectins, represent potent sensitizers to promote tumor cell killing.

Bio-Inspired Proanthocyanidins from Blueberries’ Surface Coating Prevents Red Blood Cell Agglutination on Urinary Silicon-Based Catheters

Thrombosis can cause the occlusion of implantable medical devices, leading to the rejection of the device and subsequent mortality. Thrombosis is primarily induced by red blood aggregation and coagulation. The administration of anticoagulant drugs is generally used as a treatment to avoid these processes. Adverse effects such as bleeding in the event of an anticoagulant overdose, osteoporosis associated with prolonged use, hypersensitivity, and hives have been reported. New strategies such as biomolecule surface functionalization have recently been studied to overcome these problems. In this study, we report a novel coating composed of polydopamine (PDA) and proanthocyanidins (PACs) from blueberry extract to avoid red blood aggregation in short-term use medical devices such as silicone catheters. We showed that PDA formed stable films on silicone surfaces and PACs could be immobilized on PDA layers using laccase as a catalyst. The PDA–PACs films decreased surface hydrophilicity, increased surface roughness, and decreased plasma protein adsorption. The films were stable in phosphate buffer saline (PBS) and cell culture media. Furthermore, red blood cell adsorption and aggregation decreased. These effects are attributed to changes in the membrane fluidity that influences adhesion, the steric hindrance of the layers, and the low adsorption of plasma proteins on the PAC layer.

Molecular characterization and expression profiling of tandem-repeat galectin-8 from red-spotted grouper (Epinephelus akaara): Potential antibacterial, antiviral, and wound healing activities

Galectin-8 is a typical β-galactoside binding lectin, which primarily functions as a pattern recognition receptor and/or danger receptor that is engaged in pathogen recognition by the host innate immune system. Although several fish galectins have been identified, the role of galectin-8 in teleost immunity is still not fully understood. In this study, molecular, transcriptional, and immune-related functions of galectin-8 (EaGal8) from red-spotted grouper (Epinephelus akaara) were analyzed. The open reading frame of EaGal8 comprised 960 bp encoding 319 amino acids of a ∼35 kDa protein, composed of the N- and C-terminal carbohydrate recognition domains joined by a short hinge peptide. Phylogenetic analysis revealed that EaGal8 was closely related to the Epinephelus lanceolatus galectin-8-like protein. Although EaGal8 showed ubiquitous tissue expression, the highest expression level was observed in the blood. Immunostimulants, including lipopolysaccharide, poly(I:C), and nervous necrosis virus, significantly upregulated the EaGal8 transcription level in a time-dependent manner (p < 0.05). Furthermore, recombinant EaGal8 (rEaGal8) showed a binding affinity toward seven different carbohydrates in a concentration-dependent manner. In addition, rEaGal8 caused strong agglutination of fish red blood cells and several gram-positive and gram-negative bacteria, including Streptococcus iniae, Streptococcus parauberis, Lactococcus garvieae, Escherichia coli, Edwardsiella tarda, Vibrio alginolyticus, Vibrio parahaemolyticus, and Pseudomonas aeruginosa. For the first time in teleosts, we report the wound healing ability of galectin-8 in this study. At low concentrations, rEaGal8 showed potential wound healing responses in FHM cells, in vitro. Thus, this study reinforces the role of EaGal8 in innate immune responses against bacterial and viral infections and wound healing in red-spotted grouper.

An Automated Method for Blood Type Determination by Red Blood Cell Agglutination Assay

An Automated Method for Blood Type Determination by Red Blood Cell Agglutination Assay

Studies on Lectin Mediated Agglutination Reaction on Red Blood Cell Surface Antigens Using Hot and Cold Water Plant Extracts

Lectin has various physiological roles in cell agglutination, based on their carbohydrate-binding properties, plant lectins are widely used for the detection, segregation, and characterization of glycoconjugates. Rhesus (Rh) factor is a protein that is inherited and found on the surface of red blood cells. If the surface protein is present, the RBC is Rh positive; otherwise, it is Rh-negative in nature. In this paper, we use agglutination reactions to investigate the effect of different cold and hot water extracted plants on RBC antigens as an alternative to commercial monoclonal antibodies. Extensive research on the sequence homology and 3-D structure of various plant lectins suggests that they have been conserved throughout evolution and may play important physiological roles that are still unknown.

P06 Waldenström macroglobulinemia – beware the label of primary Raynaud’s Phenomenon

P06 Waldenström macroglobulinemia – beware the label of primary Raynaud’s Phenomenon

The Hemostatic and Wound Healing Effect of Chitosan Following Debridement of Chronic Ulcers

Chitosan has been proven to be helpful in wound care as a hemostatic agent. The hemostatic effect is due to the positively charged chitosan interacting with negatively charged red blood cell membranes, initiating the agglutination of red blood cells and platelets. This promotes the activation of thrombin, which activates the clotting pathway, leading to thrombus formation. Based on the properties of chitosan as a rapidly acting hemostatic agent, the authors sought to determine if a chitosan gelling fiber wound dressing could control bleeding of freshly debrided wounds. The effect of the chitosan dressing on overall healing and patient and provider satisfaction was also evaluated. Wounds of any etiology requiring sharp debridement in patients older than 18 years who were capable of consent were eligible. Wounds were sharply debrided by curettage, scalpel, electrosurgery, or a combination of methods. A chitosan dressing was applied to the freshly debrided wound with gentle pressure. The time from application to hemostasis as assessed by non-progression of blood pattern was measured. Other outcome measures also included digital photography, wound surface area, numerical pain scores, and Photographic Wound Assessment Tool (PWAT) scores. Patient and provider satisfaction were measured. Twenty patients with a variety of etiologies and ulcer types were evaluated. After debridement, wound bleeding was rated as mild (n=9), moderate (n=9), or severe (n=2). The mean time to hemostasis was 75 seconds ± 41 SD (range, 28-221 seconds). In 1 week, the mean wound area decreased from 6.9 cm2 ± 7.8 to 6.2 cm2 ± 7.9 and mean PWAT scores decreased from 17.7 ± 4.9 to 11.4 ± 5.0 (lower score indicates wound healing). Pain scores associated with wound debridement were reduced in all but 1 patient evaluated at week 1. Overall, the rating scores from the Patient Reported Acceptance Questionnaire (PRAQ) and Provider Acceptance Questionnaire (PAQ) developed by this research group were high. The mean total PRAQ score was 30.5 ± 3.9 out of 35 (35 being most satisfied). The PAQ score was 15 out of 15 for all but 1 patient (15 being most satisfied). The chitosan gelling fiber wound dressing was simple to use and rapidly promoted hemostasis in fresh sharply debrided wounds. It was safe and easy to use in an outpatient setting and was highly rated by the patients.

Characterization of red blood cells agglutination based on stochastic collision electrochemical technique

Hemagglutination assays are important for blood group typing, alloantibodies screening and biomarkers detecting. Simple and sensitive approach for both detecting and quantifying hemagglutination is in need of development. In this research, the specific agglutination of red blood cells (RBCs) was characterized by stochastic collision events of RBCs on a Pt ultramicroelectrode (UME), extending the collision electrochemical technique to the biological reaction analysis. The collision and irreversible adsorption of RBCs blocked the oxidation flux on the UME and resulted in the “staircase” current drop in the chronoamperometry curve. The concentration, mean diameter and coverage ratio of RBCs were determined. Optical tracking of RBCs collision events verified the size and landing position dependence of RBCs to their collision step magnitudes. Adding specific antibodies to the RBCs, antibodies would connect RBCs together as a “bond” and form RBCs clusters. Such agglutination manifested itself by the decrease in collision frequency and the remarkable increase in the step current magnitude. Also, the agglutination extent could be roughly estimated from the largest step height or quantitatively determined from the decrease ratio of collision frequency. This work shows the reliability and sensitivity of collision electrochemical technique for hemagglutination analysis and builds the foundation for its further application on the blood-grouping sensors and immunoassays of infectious diseases.

Glycosylation reduces the glycan-independent immunomodulatory effect of recombinant Orysata lectin in Drosophila S2 cells

Several plant lectins, or carbohydrate-binding proteins, interact with glycan moieties on the surface of immune cells, thereby influencing the immune response of these cells. Orysata, a mannose-binding lectin from rice, has been reported to exert immunomodulatory activities on insect cells. While the natural lectin is non-glycosylated, recombinant Orysata produced in the yeast Pichia pastoris (YOry) is modified with a hyper-mannosylated N-glycan. Since it is unclear whether this glycosylation can affect the YOry activity, non-glycosylated rOrysata was produced in Escherichia coli (BOry). In a comparative analysis, both recombinant Orysata proteins were tested for their carbohydrate specificity on a glycan array, followed by the investigation of the carbohydrate-dependent agglutination of red blood cells (RBCs) and the carbohydrate-independent immune responses in Drosophila melanogaster S2 cells. Although YOry and BOry showed a similar carbohydrate-binding profiles, lower concentration of BOry were sufficient for the agglutination of RBCs and BOry induced stronger immune responses in S2 cells. The data are discussed in relation to different hypotheses explaining the weaker responses of glycosylated YOry. In conclusion, these observations contribute to the understanding how post-translational modification can affect protein function, and provide guidance in the selection of the proper expression system for the recombinant production of lectins.

Pigmented Corn Varieties as Functional Ingredients for Gluten-Free Products.

Oxidative stress, one among the several factors responsible for the gluten toxicity in celiac disease, together with inflammation and duodenal mucosal injury, are only partially reduced by the gluten-free diet. Thanks to their phenolic profile, the pigmented varieties of corn could be an interesting source of dietary antioxidants for the formulation of new gluten-free ingredients. The aim of this research was: (1) to characterize the phenolic profile and the associated antioxidant properties of corn samples with different pigmentation, using spectrophotometric and chromatographic techniques and (2) to assess the stability of anthocyanins during the gastro-intestinal digestion. The pigmented varieties showed a significantly higher content of polyphenols compared to the common yellow varieties and, as a consequence, a higher antioxidant activity. Although corn is among the cereals most frequently used in gluten-free products, it can produce an inflammatory response in some celiac patients. Therefore, after the chemical characterization, the safety of the pigmented varieties for celiac patients was confirmed using different in vitro models (cell agglutination test and the measure of transepithelial electrical resistance). Although in vivo studies are necessary, the data collected in this study underline that the pigmented corn could have a role in reducing the oxidative stress at the intestinal level in celiac subjects.

In vitro Assessment of Cytotoxic Activity of Bioactive Peptides from Momordica dioica and Solanum trilobatum against Human Colon Cancer Cells

Plant peptides have gained attention in the medicinal field due to their high anti-microbial and anti-cancer properties. Various plant sources are being used to extract proteins and peptides to be used as a cure for a variety of diseases. The latest studies show that plant peptides are effective in the treatment of cancer due to their ability to preferentially bind to the receptors or membranes of cancer cells leading to tumour growth inhibition, cytotoxicity, decreased proliferation, and apoptosis. The peptides get internalized into the cells causing cancer cell agglutination and aggregation. In addition to acting as therapeutic agents, plant peptides are also used for the targeting of drugs specific to cancer cells. In this study, bioactive peptides were isolated from the seeds of Momordica dioica and leaves of Solanum trilobatum. They were screened and identified using HPLC and MALDI-TOF techniques. In this study, apoptosis was analyzed by the Hoechst 33342 staining method which detected the presence of condensed pycnotic nuclei in apoptotic COLO320DM and COLO205 colon cancer cellsat the maximum concentrations of 150 and 175 µg/mL of plant peptides.Further DCFH-DA staining indicated the intracellular ROS production in the treated COLO320DM and COLO205 colon cancer cells. Thus, the isolated bioactive plant peptides can be formulated towards the development of effective anticancer drugs for the treatment of colon cancer in humans.

The lectin Orysata induces phosphatase-mediated and carbohydrate-independent aggregation of insect cells

Lectins, or carbohydrate-binding proteins, can cause agglutination of particular cells. This process is mediated by the interaction of the carbohydrate-binding domain with sugar structures on the cell surface, and this binding can be inhibited by pre-incubation of the lectin with its specific sugars. However, when incubated with insect cells, Orysata, a mannose-binding lectin from rice, caused aggregation of the cells, independent from carbohydrate binding activity. This phenomenon was observed for multiple insect cell lines, confirming the robustness of this phenotype. While the carbohydrate-dependent agglutination of red blood cells happens within minutes, the carbohydrate-independent aggregation of insect cells requires longer incubation times. Further analysis with the galactose-binding lectins SSA and Jacalin, validated the robustness of this lectin-induced, carbohydrate-independent aggregation in different insect cell lines. Since proteomic analysis revealed no changes in the proteome after treatment with the lectins, this cell aggregation is likely caused by the (in) activation or re-organization of the existing surface proteins. The use of inhibitors of phosphorylation and dephosphorylation, staurosporine (STS) and a phosphatase inhibitor (PPI) cocktail, pointed to dephosphorylation as a key mechanism in the lectin-induced, carbohydrate-independent aggregation of insect cells. Similar to contact inhibition, cell proliferation in cell aggregates was decreased. Analysis of the marker for cell proliferation, cyclin E, confirmed that aggregated cells enter a quiescent state. The current data offer a new perspective on the mechanism by which lectins execute their activities, specifically through lectin-induced phosphatase-mediated cell aggregation and proliferation inhibition, independent from their carbohydrate-binding activity.

Cold agglutinin-induced hemolytic anemia during room temperature fluid resuscitation: a case report

BackgroundCold agglutinin disease can cause the agglutination of red blood cells and hemolytic anemia due to cold temperature. Herein, we report a case of progressive hemolytic anemia due to cold agglutinin disease during fluid resuscitation and in the absence of exposure to cold.Case presentationA 71-year-old Japanese man was admitted to the emergency department with signs of hypotension and disturbed consciousness. He was diagnosed with diabetic ketoacidosis, and treatment with fluid resuscitation and insulin infusion was initiated. Laboratory test results obtained the following day indicated hemolytic anemia. On day 5 after admission, red blood cell agglutination was detected, and the patient was diagnosed with cold agglutinin disease.ConclusionsCold agglutinin disease should be considered in the differential diagnosis of progressive hemolytic anemia during fluid resuscitation, even if the solution is at room temperature.

Perfusion bioreactor enabled fluid-derived shear stress conditions for novel bone metastatic prostate cancer testbed

Metastatic prostate cancer spreads preferentially to the bone, causing skeletal complications associated with significant morbidity and a poor prognosis, despite current therapeutic approaches. Hence, it is imperative to understand the complex metastatic cascade to develop therapeutic interventions for treating metastatic prostate cancer. Increasing evidence suggests the synergistic role of biochemical and biophysical cues in cancer progression at metastases. However, the mechanism underlying the crosstalk between interstitial flow-induced mechanical stimuli and prostate cancer progression at the bone microenvironment remains poorly understood. To this end, we have developed a three-dimensional (3D) in vitro dynamic model of prostate cancer bone metastasis using perfusion bioreactor and compared our results with static conditions to delineate the role of flow-induced shear stress on prostate cancer progression at metastases. We observed an increase in human mesenchymal stem cell (hMSCs) proliferation and differentiation rate under the dynamic culture. The hMSCs form cell agglutinates under static culture, whereas the hMSCs exhibited a directional alignment with broad and flattened morphology under dynamic culture. Further, the expression of mesenchymal to epithelial transition biomarkers is increased in bone metastasized prostate cancer models, and large changes are observed in the cellular and tumoroid morphologies under dynamic culture. Evaluation of cell adhesion proteins indicated that the altered cancer cell morphologies resulted from the constant force pulling due to increased E-cadherin and phosphorylated focal adhesion kinase proteins under shear stress. Overall, we report a successful 3D in vitro dynamic model to recapitulate bone metastatic prostate cancer behavior under dynamic conditions.

A haemagglutination test for rapid detection of antibodies to SARS-CoV-2

Serological detection of antibodies to SARS-CoV-2 is essential for establishing rates of seroconversion in populations, and for seeking evidence for a level of antibody that may be protective against COVID-19 disease. Several high-performance commercial tests have been described, but these require centralised laboratory facilities that are comparatively expensive, and therefore not available universally. Red cell agglutination tests do not require special equipment, are read by eye, have short development times, low cost and can be applied at the Point of Care. Here we describe a quantitative Haemagglutination test (HAT) for the detection of antibodies to the receptor binding domain of the SARS-CoV-2 spike protein. The HAT has a sensitivity of 90% and specificity of 99% for detection of antibodies after a PCR diagnosed infection. We will supply aliquots of the test reagent sufficient for ten thousand test wells free of charge to qualified research groups anywhere in the world.

Lectin and Its Function

Lectins are carbohydrate-binding proteins that are profoundly particular for sugar bunches that are portion of other particles and so cause agglutination of specific cells or precipitation of glycoconjugates and polysaccharides. Lectins have a part in acknowledgment on the cellular and atomic level and play various parts in natural acknowledgment marvels including cells, carbohydrates, and proteins. Lectins too intervene connection and authoritative of microscopic organisms, infections, and organisms to their aiming targets.